Integration of BpMADS4 on various linkage groups improves the utilization of the rapid cycle breeding system in apple

Rapid cycle breeding in apple is a new approach for the rapid introgression of agronomically relevant traits (e.g. disease resistances) from wild apple species into domestic apple cultivars (Malus × domestica Borkh.). This technique drastically shortens the long‐lasting juvenile phase of apple. The utilization of early‐flowering apple lines overexpressing the BpMADS4 gene of the European silver birch (Betula pendula Roth.) in hybridization resulted in one breeding cycle per year. Aiming for the selection of non‐transgenic null segregants at the end of the breeding process, the flower‐inducing transgene and the gene of interest (e.g. resistance gene) that will be introgressed by hybridization need to be located on different chromosomes. To improve the flexibility of the existing approach in apple, this study was focused on the development and characterization of eleven additional BpMADS4 overexpressing lines of four different apple cultivars. In nine lines, the flowering gene was mapped to different linkage groups. The differences in introgressed T‐DNA sequences and plant genome deletions post‐transformation highlighted the unique molecular character of each line. However, transgenic lines demonstrated no significant differences in flower organ development and pollen functionality compared with non‐transgenic plants. Hybridization studies using pollen from the fire blight‐resistant wild species accession Malus fusca MAL0045 and the apple scab‐resistant cultivar ‘Regia’ indicated that BpMADS4 introgression had no significant effect on the breeding value of each transgenic line.     

Prenatal diagnosis in a mentally retarded woman with mosaic ring chromosome 18

We present a pregnant woman with mental retardation and mosaic for ring 18 referred for prenatal diagnosis. Major clinical features included short stature with clinodactyly in feet, foot deformity and club feet, hypotonia, kyphosis, and absence of breast development, low set ears, high arched palate, dental decay and speech disorder. Prenatal diagnosis was carried. Using amniocentesis. The fetus had a normal karyotype described as 46,XX. The fetus was evaluated for clinical features after delivery; she was healthy with no abnormal clinical characterizations.     

红掌茎段侧芽离体快繁技术研究

以红掌嫩茎为外植体,诱导侧芽萌发,并进行增殖和生根培养,研究不同生长调节剂浓度配比对茎段侧芽萌发、增殖、无菌苗生根的影响以及增殖培养过程中愈伤组织的抑制等因素。结果表明,侧芽诱导的适宜培养基为MS+6-BA 2.0 mg/L+NAA 0.5 mg/L,萌发率达87.5%;最适增殖培养基为MS+6-BA 0.8 mg/L+NAA 0.2 mg/L+VB2 8.0 mg/L,增殖系数3.8;最适生根培养基为1/2MS+NAA 0.5 mg/L,生根率98%;在增殖培养基中添加适量VB2能较好地抑制愈伤组织的生成,防止愈伤组分织分化形成芽,从而达到以芽繁芽的目的。     

Characterization and functional activity of murine monoclonal antibodies specific for α1,6-glucan chain of Helicobacter pylori lipopolysaccharide

Background: The outer core region of H. pylori lipopolysaccharide (LPS) contains α1,6‐glucan previously shown to contribute to colonizing efficiency of a mouse stomach. The aim of the present study was to generate monoclonal antibodies (mAbs) specific for α1,6‐glucan and characterize their binding properties and functional activity. Materials and Methods: BALB/c mice were injected intraperitoneally with 10⁸ formalin‐fixed H. pylori O:3 0826::Kan cells 3× over 56 days to achieve significant titer. Anti‐α1,6‐glucan‐producing hybridomas were screened by indirect ELISA using purified H. pylori O:3 0826::Kan LPS. One clone, 1C4F9, was selected for further characterization. The specificities of mAbs were determined by indirect and inhibition ELISA using structurally defined H. pylori LPS and synthetic oligosaccharides, and whole‐cell indirect ELISA (WCE) of clinical isolates. They were further characterized by indirect immunofluorescent (IF) microscopy and their functional activity in vitro determined by serum bactericidal assays against wild‐type and mutant strains of H. pylori. Results: The generated anti‐α1,6‐glucan IgM, 1C4F9, has demonstrated an excellent specificity for the glucan chain containing 5 to 6 α1,6‐linked glucose residues and showed surface accessibility by IF microscopy with H. pylori cells adherent to gastric adenocarcinoma cells monolayers. Of 38 isolates from Chile, 17 strains reacted with antiglucan mAbs in WCE (OD₄₅₀ ≥ 0.2). Bactericidal activity was observed against selective wild‐type and mutant H. pylori strains exhibiting OD₄₅₀ values of ≥0.45 in WCE. Conclusions: Anti‐α1,6‐glucan mAbs could have potential application in typing and surveillance of H. pylori isolates as well as offer insights into structural requirements for the development of LPS‐based vaccine against H. pylori infections.     

Cloning, expression and purification of extracellular serine protease Esp, a biofilm-degrading enzyme, from Staphylococcus epidermidis

Aims: Staphylococcus epidermidis Esp, an extracellular serine protease, inhibits Staphylococcus aureus biofilm formation and nasal colonization. To further expand the biotechnological applications of Esp, we developed a highly efficient and economic method for the purification of recombinant Esp based on a Brevibacillus choshinensis expression–secretion system. Methods and Results: The esp gene was fused with the N‐terminal Sec‐dependent signal sequence of the B. choshinensis cell wall protein and a C‐terminal hexa‐histidine‐tag gene. The recombinant Esp was expressed and secreted into the optimized medium as an immature form and subsequently activated by thermolysin. The mature Esp was easily purified by a single purification step using nickel affinity chromatography and showed proteolytic activity as well as Staph. aureus biofilm destruction activity. Conclusions: The purification yield of the developed extracellular production system was 5 mg recombinant mature Esp per 20‐ml culture, which was much higher than that of an intracellular production system in Escherichia coli (3 mg recombinant Esp per 1‐l culture). Significance and Impact of the Study: Our findings will be a powerful tool for the production and purification of recombinant Esp and also applicable to a large variety of recombinant proteins used for basic researches and biotechnological applications.     

The challenge to quantify Listeria monocytogenes--a model leading to new aspects in molecular biological food pathogen detection

In this work, we discuss the latest insights concerning advantages and disadvantages and the nature of microbiological and molecular methods for quantitative food pathogen detection. The assessment of molecular methods must be brought on a basis that considers the nature of molecular methods and their underlying mechanism. A potential approach to setting up the development, validation and structure of an analytical chain is presented based on quantitative real-time PCR (qPCR). This is analysed exemplary on the basis of recent work using the model organism Listeria monocytogenes. Several prerequisites for successful quantitative detection of this pathogen will be discussed. In particular, sample preparation, controls for all methodical steps and the validation of the core assay qPCR are addressed, which constitute the basis for a reliable analytical detection chain for molecular biological pathogen detection from food. Microbiological methods are analysed based on growth of the single cell, which is the fundament of these traditional methods.     

利用与PKD1紧密连锁的微卫星多态性对黑龙江省汉族人群进行ADPKD的症前诊断

目的：利用与PKD1紧密连锁的微卫星的多态性对黑龙江省汉族人群常染色体显性遗传性多裳肾病（autosomal dominant polycystic diseaseADPKD）家系成员进行症状前诊断。方法：扩增PKD1基因内部（KG8）及基因两侧（AC2．5、CW4）共三个微卫星遗传标记,并采用聚丙烯酰胺凝胶电泳进行检测,银染,最后进行家系连锁分析。结果：家系1中一名男孩携带了PKD1突变基因,处于发病前期。结论：通过连锁分析ADPKD进行连锁分析能高效、快捷、准确地进行早期诊断.     

肿瘤标志物联合检测在乳腺癌早期诊断中的应用

目的：探讨肿瘤标志物癌胚抗原（CEA）、糖链抗原19-9（CA19-9）、糖链抗原15-3（CA15-3）联合检测在乳腺癌早期诊断中的应用价值。方法：检测87例乳腺癌患者,55例乳腺良性肿瘤患者和35例健康人血清中CEA、CA19-9、CA153等肿瘤标志物的水平及3种标志物不同组合对乳腺癌的阳性检出率。结果：乳腺癌患者3种肿瘤标志物显著高于正常对照组及乳腺良性肿瘤组（P〈0.01）。3项标志物不同组合对不同分期乳腺癌检出的敏感性均高于单项标志物。其中CEA＋CA199＋CA153组合的检出敏感性较其他组合均高,特别是对早期患者检出率明显提高。结论：CEA＋CA199＋CA153联合检测能提高乳腺癌的早期诊断率。